野生型p53基因对卵巢癌细胞株生长及凋亡的影响

Inhibition and apoptosis of the control of ovarian cancer cell line induced by introduction of recombinant wild Type p53 gene

目的 探讨野生型 p5 3基因对卵巢癌细胞生长抑制及凋亡的作用 ,为卵巢癌的基因治疗提供实验依据。方法 构建野生型 p5 3基因重组腺病毒载体、体外转染卵巢癌细胞株CaOV3;应用生化染色、免疫组化、聚合酶链技术 ,检测外源基因的转染率及表达效果 ;应用细胞计数、MTT法 (四甲基偶氮唑盐微量酶反应比色法 )、流式细胞术和TUNEL技术 (TDT -mediateddUTP -biotinendlabeling) ,检测细胞生长及凋亡的情况。 结果 野生型p5 3基因重组腺病毒载体可以有效地转染卵巢癌细胞株CaOV3,转染后的CaOV3细胞内可以检测到 p5 3基因的cDNA及p5 3蛋白的表达 ;转染后的CaOV3细胞生长受到明显抑制 ,6 3%的细胞生长停滞于G0 /G1期 ,4 0 %～ 5 0 %的细胞TUNEL呈阳性。结论 野生型 p5 3基因的导入 ,可抑制卵巢癌细胞的生长并诱导细胞凋亡。野生型 p5 3基因导入可能成为今后临床基因治疗卵巢癌的可行性方法之一。

Objective To study the role of inhibition on the growth and apoptosis of ovarian cancer cell line by the introduction of wild-Type p53 gene and investigate the mechanism of gene therapy for ovarian cancer. Methods The recombinant p53 gene adenovirus vector was constructed and transfected into the ovarian cancer cell line CaOV3. The transfecting efficiency of p53 gene and expression of p53 protein were detected by biochemistry stain, immunohistochemical analysis, PCR technique and agarose gel electrophoresis. The inhibition on the growth and apoptosis of CaOV3 were tested by cell counting, flow cytometry, MTT (Tetrazolium salts colorimetric assay), TUNEL technique (terminal deoxynucleotidyl transferase mediated dUTP nick end labeling). Results The constructed recombinant p53 gene adenovirus vector could transfect into the CaOV3 cells efficiently,p53 cDNA and protein expression could be founded in the infected cells. The proliferation of transfected CaOV3 was inhibited evidently. 63% cells arrested at G 0/G 1 phases of cell cycle,40%～50% cells showed positive TUNEL staining. Conclusion The introduction of recombinant wild type p53 gene can suppress CaOV3 cells proliferation and cause apoptosis efficiently. It might be a suitable method on gene treatment for ovarian cancer.